It is well known that when a foreign substance such as a bacterium or virus enters a human or animal body, production of antibodies to combat the infection is stimulated by the presence of one or more antigens. Antigens related to invading organisms comprise a foreign substance that comes from the invading organism, such as a piece of bacterium or virus. More generally, an antigen is any substance capable of triggering an immune response. Certain specialized cells of the body contact an antigen and produce antibodies specifically designed for that antigen. When released in the body, such antibodies identify and bind to the antigen, thereby combating the infection. Antibodies are highly specific and will generally bind only to the antigen which stimulated their creation.
When a person has been infected with certain diseases, that person's blood will often contain measurable levels of antigen specific to that disease. To determine whether such an infection is present, an immunodiagnostic test is performed using a sample of the patient's blood. The sample is mixed with a solution known to contain antibodies specific to a certain disease or condition. If an antigen-antibody reaction occurs, the test result is positive, and the antigen is detected. Such a test is typically reversible, i.e., a solution or reagent known to contain a certain antigen can be used to determine whether or not the corresponding antibody is present in a sample. However, the antigen-antibody reaction occurs on a microscopic level and is not readily observable. Thus, all known immunodiagnostic tests provide some type of means for indicating that the antigen-antibody reaction has occurred.
A variety of techniques have been used to detect antigen-antibody reactions. The principal techniques presently in use are enzyme immunoassay, immunofluorescence, and radioimmunoassay. In typical enzyme immunoassay procedures, the antigen-antibody reaction is detected by the formation, by an enzyme, of a colored product from a colorless substrate. Immunofluorescence techniques indicate that a reaction has occurred by emission of small quantities of light which must generally be observed microscopically. Radioimmunoassay utilizes radioactive labeling substances so that occurrence of the antigen-antibody reaction is measured by the presence or absence of small amounts of radioactivity. These known methods are reliable but are slow and tedious.
Recently several types of electrical immunoassay techniques have been developed. One such technique utilizes field effect transistors coated with a layer of antibody in the gate region. If an antigen-antibody reaction occurs, the charge concentration of the transistor changes. Examples of this type of system are given in Schenck U.S. Pat. No. 4,238,757, issued Dec. 9, 1980; Guckel U.S. Pat. No. 4,180,771, issued Dec. 25, 1981; Malmros U.S. Pat. Nos. 4,334,880, issued Jun. 15, 1982 and 4,444,892, issued Apr. 24, 1984, and Japanese Patent Publication No. 60-29658. In another system, a body fluid containing an analyte to be detected is deposited onto a surface which has been coated with a reagent that binds specifically to the analyte, so that a binding reaction takes place. A tagged reagent is then added which reacts with the analyte-reagent complex or with the reagent to change the electrical reactance of the surface. See Ebersole U.S. Pat. No. 4,219,335, issued Aug. 26, 1980.
Several other methods have been proposed for measuring immunologic reactions electrically. A voltametric immunoassay can be carried by labeling one immunoreactant with an electroactive substance. Pace U.S. Pat. No. 4,233,144, issued Nov. 11, 1980, is illustrative of one such technique. Another method involves sandwiching an antigen-antibody layer between two conductive layers and measuring the electrical capacitance of the resulting laminate. Giaever U.S. Pat. No. 4,054,646, issued Oct. 18, 1977, describes such a method. A further type of capacitance-measuring system includes a pair of electrodes coated with a substrate and immersed in a medium containing a material which specifically binds with the substrate, as described in Arwin U.S. Pat. No. 4,072,576. A further method combines change effect signal detection with an enzyme immunoassay technique. Such a method is disclosed by Gibbons U.S. Pat. No. 4,287,300, issued Sep. 1, 1981. The foregoing electrical methods have, however, failed to provide medical practitioners and laboratories with a simple, fast, sensitive, inexpensive and easy-to-use method of performing an immunodiagnostic test.
One aspect of the present invention involves the use of antigen or antibody-labeled colloidal gold particles. In general, "colloidal gold" refers to a suspension of fine gold particles in water or aqueous solution. Preparation of such particles is disclosed by DeMey, et al. U.S. Pat. No. 4,446,238, issued May 1, 1984, and DeMey, et al. U.S. Pat. No. 4,420,558, issued Dec. 13, 1983. The entire contents of both such DeMey patents are incorporated herein by reference. Such colloidal gold preparations have been previously used in immunodiagnostic tests wherein the results are determined optically by observing small amounts of light reflected as a result of the antigen-antibody reaction. The foregoing patents to DeMey disclose a bright field light method of the foregoing type. Silver enhancement has been previously used as a means for staining gold particles. The present invention advantageously employs colloidal gold, optionally with silver enhancement, in a new immunodiagnostic method.